AZD-8542

Mesenchyme homeobox-2 (MEOX2)-mediated regulation of glioma-associated oncogene-1 (GLI1) has been associated with poor overall survival, conferring chemoresistance in lung cancer. However, the role of MEOX2/GLI1 in resistance to epidermal growth factor receptor-tyrosine kinase inhibitors (EGFR-TKIs)-based therapy remains unexplored in human lung cancer.

Functional assays using genetic silencing strategy by short hairpin RNAs, as well as cytotoxic (tetrazolium dye MTT) and clonogenic assays, were performed to evaluate MEOX2/GLI1-induced malignancy capacity in lung cancer cells. Further analysis performed includes western blot, qPCR and ChIP-qPCR assays to identify whether MEOX2/GLI1 promote EGFR/AKT/ERK activation, as well as EGFR overexpression through epigenetic mechanisms. Finally, preclinical tumour progression in vivo and progression-free disease interval analyses in patients treated with EGFR-TKI were included.

Overexpressed MEOX2/GLI1 in both EGFR wild-type and EGFR/KRAS-mutated lung cancer cells were detected and involved in the activation/expression of EGFR/AKT/ERK biomarkers. In addition, MEOX2/GLI1 was shown to be involved in the increased proliferation of tumour cells and resistance capacity to cisplatin, EGFR-TKIs (erlotinib and AZD9291 'osimertinib'), AZD8542-SMO, and AZD6244-MEKK1/2. In addition, we identified that MEOX2/GLI1 promote lung tumour cells progression in vivo and are clinically associated with poorer progression-free disease intervals. Finally, both MEOX2 and GLI1 were detected to be epigenetically involved in EGFR expression by reducing both repressive markers polycomb-EZH2 and histone H3K27me3, but, particularly, increasing an activated histone profile H3K27Ac/H3K4me3 at EGFR-gene enhancer-promoter sequences that probably representing a novel EGFR-TKI-based therapy resistance mechanism.

MEOX2/GLI1 promote resistance to cisplatin and EGFR-TKI-based therapy in lung cancer cells, modulating EGFR/AKT/ERK signalling pathway activation, as well as inducing an aberrant epigenetic modulation of the EGFR-gene expression in human lung cancer.

Aberrant hedgehog (Hh) pathway signaling is implicated in multiple cancer types and targeting the Smoothened (SMO) receptor, a key protein of the Hh pathway, has proven effective in treating metastasized basal cell carcinoma. Our lead optimization effort focused on a series of heteroarylamides. We observed that a methyl substitution ortho to the heteroaryl groups on an aniline core significantly improved the potency of this series of compounds. These findings predated the availability of SMO crystal structure in 2013. Here we retrospectively applied quantum mechanics calculations to demonstrate the o-Me substitution favors the bioactive conformation by inducing a dihedral twist between the heteroaryl rings and the core aniline. The o-Me also makes favorable hydrophobic interactions with key residue side chains in the binding pocket. From this effort, two compounds (AZD8542 and AZD7254) showed excellent pharmacokinetics across multiple preclinical species and demonstrated in vivo activity in abrogating the Hh paracrine pathway as well as anti- tumor effects.